RAPD分析与ITS序列分析在拟茎点霉分类鉴定上的应用

APPLICATION OF RAPD AND ITS REGION SEQUENCE ANALYSES ON CLASSIFICATION AND IDENTIFICATION OF PHOMOPSIS

采用随机扩增多态性DNA(RAPD)技术和核糖体RNA基因(rDNA)转录间隔区(ITS)序列分析对22种拟茎点霉共34个菌株进行了系统发育研究。RAPD分析构建的UPGMA聚类图所反映的种间、种内关系与形态学分类结果基本一致,可以清楚地将分自7科寄主植物上的不同的种分别区分开来,但分自同科或同属寄主植物上的不同的种并不具有相近的亲缘关系。ITS序列分析结果不支持将Phomopsis mangiferae Ahmad和P. cytosporella Penz. et Sacc. 合并为同一个种的观点,同时还显示出P. mangiferae与P. psidii de Camara的亲缘关系非常近, 可能是异名同物;而为害木棉叶的拟茎点霉与杨梅枝枯病菌P. myricae Y.J.Huang et P.K.Chi之间的碱基差异亦属于种下不同菌株间的正常差别范围,很可能就是同一个种。对相同的供试菌株两技术所反映的亲缘关系趋势相同,表明两技术用于拟茎点霉的亲缘关系分析和种类鉴定均是可行的。

Random amplified polymorphic DNA (RAPD) markers and the DNA sequences of ribosomal DNA internal transcribed spacer (ITS) were used to analyze the relationships of 34 isolates of 22 Phomopsis species. The relationships of interspecies or intraspecies, which were showed in the dendrogram produced by UPGMA based on RAPD data, were nearly consistent with the results of morphological study. Different species isolated from plants of 7 families could be clearly distinguished in the dendrogram. But there were no close relationships among species isolated from the same family or genus of plants. ITS sequence analysis did not support P. mangiferae Ahmad and P. cytosporella Penz. et Sacc. had close relationship, whereas indicated P. mangiferae was closely related to P. psidii de Camara, implying P. psidii may be a synonym of P. mangiferae. In addition, the absolute distance between one isolate of Phomopsis durionis H. Syd (host is Bombax malabaricum D C.) and P. myricae Y.J.Huang et P.K.Chi fell within the normal range of different isolates of a single species. This suggests that they are probably conspecific. Results of RAPD analysis are consistent with those of ITS sequence analysis for the same isolates tested. Consequently both methods can be independently applied to classification and identification of Phomopsis.